Herpes simplex virus (HSV-1) is the cause of important human illnesses including disseminated disease in the newborn, cold sores, genital lesions, non-epidemic encephalitis in adults, retinitis and diseases in the immunosuppressed. The goal of the proposed project is to study assembly of the virus capsid with the aim of identifying novel targets against which small molecule anti-HSV-1 therapeutics might be directed. Studies will be carried out with HSV-1, but it is expected that the results will illuminate the steps of capsid formation as they occur in other herpesviruses as well. Like all herpesviruses, HSV-1 consists of an icosahedral capsid surrounded by a membrane envelope. The capsid contains the virus DNA. Assembly of progeny virions begins with formation of the capsid, a process that takes place in the infected cell nucleus. A DNA-free capsid shell is first formed and later packaged with DNA. The proposed studies are focused on the portal complex (a 12-mer of the UL6 gene product) through which DNA enters the capsid, and on the major capsid protein, VP5. Studies with UL6 will be devoted to testing the idea that it is involved in initiation of capsid assembly. Function of UL6 in initiation would solve the problem of how it becomes incorporated at a unique site in the capsid. Studies with VP5 are devoted to clarifying the extensive conformational re-arrangement it undergoes as it becomes incorporated into the nascent capsid and as the capsid matures. Specific goals of the project are to: (1) examine the capsid location and structure in solution of UL6, the subunit protein of the portal complex; (2) test the idea that HSV-1 capsid formation is initiated by structures composed of the scaffolding and portal proteins; (3) determine the mechanism of action of WAY-150138, a small molecule (a substituted thiourea) that inhibits HSV-1 growth in cell culture by blocking DNA encapsidation; and (4) use sensitivity to trypsin digestion to clarify the structural transformation of the HSV-1 major capsid protein, VP5, as it participates in procapsid assembly and maturation.